Imaging of oxygen by phosphorescence quenching

Abstract

A method of real-time imaging of dissolved oxygen concentration, comprising adding an oxygen-quenched phosphorescent composition to a sample, exciting phosphorescence in the composition by illuminating the sample with pulses of light, detecting phosphorescence intensity as a function of position in the sample at first and second times following exciting pulses of light, determining oxygen concentration from the phosphorescence detected at the first and second times, generating an image of the oxygen concentration as a function of position, and repeating the exciting, detecting, determining, and image generating steps to produce a series of images showing the oxygen concentration varying over time.

What is claimed is: 
     
       1. A method of imaging of dissolved oxygen concentration in a sample, comprising:
 adding to a sample an oxygen-quenched phosphorescent substance, 
 exciting phosphorescence in the phosphorescent substance by illuminating the sample with pulsed light, 
 detecting phosphorescence intensity as a function of position in the sample at first and second times following an exciting pulse of light, 
 determining oxygen concentration from the phosphorescence detected at the first and second times, 
 generating an image of the oxygen concentration as a function of position, and 
 repeating the exciting, detecting, determining, and image generating steps to produce a series of images showing the oxygen concentration over time. 
 
     
     
       2. The method of  claim 1 , wherein detecting phosphorescence intensity at the first and second times comprises detecting phosphorescence over a period starting at a first time after an exciting pulse and a period starting at a second time after an exciting pulse. 
     
     
       3. The method of  claim 2 , wherein detecting phosphorescence intensity at the first and second times comprises integrating the phosphorescence detected over the period starting at the first time and over the period starting at the second time. 
     
     
       4. The method of  claim 1 , wherein detecting phosphorescence intensity at the first and second times following an exciting pulse of light comprises detecting phosphorescence at a first time following a first exciting pulse of light and detecting phosphorescence at a second time following a second exciting pulse of light. 
     
     
       5. The method of  claim 4 , wherein the first and second exciting pulses of light are successive pulses of light in a periodic stream of pulses, and repeating the exciting step comprises illuminating the sample with further pulses in the periodic stream of pulses. 
     
     
       6. The method of  claim 5 , wherein the first and second exciting pulses of light are consecutive pulses of light in the periodic stream of pulses, repeating the detecting step comprises detecting phosphorescence intensity at the first time following a third pulse consecutively following the second exciting pulse, and repeating the determining step comprises determining oxygen concentration from the phosphorescence detected at the first time following the third pulse and at the second time following the second pulse. 
     
     
       7. The method of  claim 1 , further comprising carrying out the repeating step at least 25 times a second. 
     
     
       8. The method of  claim 1 , wherein the exciting step of the repeating step begins before the previous image generating step is completed. 
     
     
       9. The method of  claim 7  wherein the exciting step of the repeating step begins before the previous determining step is completed. 
     
     
       10. Apparatus for real-time imaging of dissolved oxygen concentration in a sample, comprising:
 a light source for exciting phosphorescence in a phosphorescent substance by illuminating the sample with pulses of light; 
 a detector for detecting phosphorescence as a function of position in the sample at first and second times following an exciting pulse of light; 
 a processor arranged to determine a difference between the phosphorescence intensity detected at the first time and the phosphorescence detected at the second time; and 
 a display unit arranged to generate an image of a function of the determined difference as a function of position; 
 wherein the detector, processor, and display unit are arranged to generate successive images at successive times. 
 
     
     
       11. The apparatus of  claim 10 , wherein the detector is arranged to detect phosphorescence over a first period starting at the first time after an exciting pulse and over a second period starting at the second time after an exciting pulse. 
     
     
       12. The apparatus of  claim 11 , wherein the detector is arranged to integrate the phosphorescence detected over the first period and to integrate the phosphorescence detected over the second period. 
     
     
       13. The apparatus of  claim 10 , wherein the detector is arranged to detect phosphorescence at the first time following a first exciting pulse of light and at the second time following a second exciting pulse of light. 
     
     
       14. The apparatus of  claim 10 , wherein the detector comprises an intensified charge-coupled device (CCD) camera, and the first and second times are controlled by gating the intensifier. 
     
     
       15. The apparatus of  claim 10 , wherein the light source is arranged to emit a periodic stream of pulses and the detector is arranged to detect phosphorescence intensity from successive pulses of light in the periodic stream. 
     
     
       16. The apparatus of  claim 15 , wherein the detector is arranged to detect phosphorescence at the first and second times following pairs of pulses in the periodic stream of pulses wherein the second pulse of a pair of pulses is the first pulse of a subsequent pair of pulses. 
     
     
       17. The apparatus of  claim 10 , wherein the detector, processor and display unit are arranged to produce a new image at least 25 times a second. 
     
     
       18. The apparatus of  claim 10 , wherein the detector is arranged to start detecting phosphorescence for a next image before the processor has completed determining the difference for the previous image. 
     
     
       19. The apparatus of  claim 10 , wherein the light source is arranged to emit light with a peak intensity at a frequency selected to excite an oxygen-quenched phosphorescent composition. 
     
     
       20. The apparatus of  claim 10 , wherein the detector is arranged to detect light with a peak frequency sensitivity selected to correspond to a peak emission frequency of an oxygen-quenched phosphorescent composition.